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Label-Free Imaging of Single Microtubule Dynamics Using Spatial Light Interference Microscopy
journal contribution
posted on 2016-12-20, 00:00 authored by Mikhail
E. Kandel, Kai Wen Teng, Paul R. Selvin, Gabriel PopescuDue to their diameter,
of only 24 nm, single microtubules are extremely
challenging to image without the use of extrinsic contrast agents.
As a result, fluorescence tagging is the common method to visualize
their motility. However, such investigation is limited by photobleaching
and phototoxicity. We experimentally demonstrate the capability of
combining label-free spatial light interference microscopy (SLIM)
with numerical processing for imaging single microtubules in a gliding
assay. SLIM combines four different intensity images to obtain the
optical path length map associated with the sample. Because of the
use of broadband fields, the sensitivity to path length is better
than 1 nm without (temporal) averaging and better than 0.1 nm upon
averaging. Our results indicate that SLIM can image the dynamics of
microtubules in a full field of view, of 200 × 200 μm2, over many hours. Modeling the microtubule transport via the diffusion-advection equation, we found that the
dispersion relation yields the standard deviation of the velocity
distribution, without the need for tracking individual tubes. Interestingly,
during a 2 h window, the microtubules begin to decelerate, at 100
pm/s2 over a 20 min period. Thus, SLIM is likely to serve
as a useful tool for understanding molecular motor activity, especially
over large time scales, where fluorescence methods are of limited
utility.
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motor activity1 nm24 nmbroadband fieldsdiffusion-advection equationSingle Microtubule Dynamicslight interference microscopy0.1 nmLabel-Free ImagingSLIMpath length mapvelocity distributiontime scalesSpatial Light Interference Microscopymicrotubule transportdispersion relation yields20 min periodpath lengthfluorescence methodsextrinsic contrast agentsintensity images2 h window
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